Evidence indicates that in addition to a genetic contribution, environmental determinants also play a role in the etiology of psychiatric disorders including schizophrenia and depression (Morozov, "Advances in Biological Psychiatry, 12, eds., S. Mendlewicz and H. Praag, Karger, New York (1983)). The hypothesis of a viral contribution is suggested by the realization that viruses can induce progressive neurological disorders associated with diverse pathological findings (Morozov, ibid., (1983); Kurstak et al., "Viruses, Immunity, and Mental Disorders", Plenum, New York (1987); ter Meulen, "Seminars in Neuroscience, 3 (1991)).
Borna disease virus (BDV) is a nonsegmented, negative-stranded (NNS) RNA virus (Briese et al., Proc. Natl. Acad. Sci., USA, 91:4362-4366 (1994); Cubitt et al., J. Virol., 68:1382-1396 (1994); de la Torre, J. Virol., 68:7669-7675 (1994); and Schneemann et al., Virol., 210:1-8 (1995)) with a nuclear site for the replication and transcription of its genome (de la Torre, supra, (1994); Schneemann et al., supra, (1995); and Cubitt et al., J. Virol., 68:1371-1381 (1994)) and the use of RNA splicing for its gene expression regulation (Cubitt et al., Virus Res., 34:69-79 (1994) and Schneider et al., J. Virol., 68:5007-5012 (1994)). These features signal BDV as the prototype of a new group of animal viruses (de la Torre, supra, (1994) and Schneemann et al., supra, (1995)).
Borna disease virus (BDV) is a noncytolytic neurotropic virus that infects a wide range of vertebrate species from birds and rodents to primates. It has a variable period of incubation and diverse pathological manifestations depending on the species, immune status and age of the host, as well as route of infection and virus strain (Ludwig et al., Prog. Med. Virol., 35:107-151 (1988); Lipkin et al., Microbial Pathogenesis, 13:167-170 (1992); Richt et al., Clin. Infect. Dis., 14:1240-1250 (1992); Koprowski et al., Curr. Topics Microbiol. Immunol., 190 (1995)).
Thus, BDV causes CNS disease in several non-human vertebrate species that is manifested by behavioral abnormalities and diverse pathologies depending on the species, age and immune status of the host, as well as route of infection and virus strain (Rott et al., in "Borna Disease", eds., H. Koprowski and I. Lipkin, Springer-Verlag, Berlin, pp17-30 (1995)). For example, heightened viral expression in limbic system structures, together with astrocytosis and neuronal degeneration within the hippocampal formation, constitute the main histopathological hallmarks of BDV infection in different animal species (Gosztonji et al., in "Borna Disease", eds., H. Koprowski and I. Lipkin, Springer-Verlag, Berlin, pp39-73 (1995) and Carbone et al., J. Virol., 65:6154-6164 (1991).
In the recently published International Application WO 96/21020, rat-derived BDV viral sequences were described for encoding rat BDV polypeptide sequences corresponding to p40, p23,gp18, p57, and BDV polymerase sequences. In addition, the application presents diagnostic and therapeutic methods for treating nervous system diseases based on the use of the rat-derived nucleic acids and encoded polypeptides. However, no human-specific sequences were identified by the authors.
The reproducible and clinically definable behavioral abnormalities accompanying BDV infection of rats and non-human primates have led to the speculation that BDV could cause similar CNS dysfunctions in humans. In support of this hypothesis are the results from cross-sectional seroepidemiological studies showing an increased prevalence of antibodies that recognize BDV-specific antigens in subjects with neuropsychiatric disorders compared to the normal healthy population (Rott et al., Science, 228:755-756 (1985); Bode et al., Lancet, ii:689 (1988); VandeWoude et al., Science, 250:1278-1281 (1990); Rott et al., Arch. Virol., 118:143-149 (1991); Bode et al., J. Med. Virol., 36:309-315 (1992); Fu et al., J. Affect. Disor., 27:61-68 (1993), for review see Bode, Curr. Top. Microbiol. Immunol., 190:101-128 (1995)). Moreover, prospective studies on acute psychiatric patients have shown a high percentage of BDV seropositives among patients with major depression (Bode et al., Arch. Virol. (Suppl), 7:159-167 (1993); Bode et al., Lancet, 343:297-298 (1994); and Bode et al., Nature Med., 1:232-236 (1995)).
Recently, using flow cytometry (FCM), BDV-specific antigens have been detected in peripheral blood monocytes (PBMC) from psychiatric patients (Bode et al., supra, (1994)). In addition, the present inventors with others have detected BDV-specific RNA sequences in such PBMC (Bode et al., supra, (1995) and Kishi et al., FEBS Letters, 364:293-297 (1995)). These findings led the present inventors to investigate the possibility of isolating infectious BDV from BDV-antigen positive human PBMC.
The present invention describes the isolation and sequence characterization of human BDV. Studies using coded PBMC samples from psychiatric patients and healthy control subjects for co-cultivation with a human oligodendroglia cell line (OL cells), led to the isolation of BDV from three hospitalized psychiatric patients, but not from any of the control subjects. The isolated virus was unequivocally identified as BDV based on the sequence identification of BDV open reading frames(ORFs) p24, p16, p56, and the putative catalytic domain of the BDV L polymerase. The sequence analysis obtained with the methods and compositions of this invention indicate that BDV human isolates are genetically very closely related to BDV from naturally infected animals of different species. These results further indicate that BDV could be one of the environmental factors contributing to the pathophysiology of neuropsychiatric disorders whose etiology remains elusive.
The present invention describes the detection of novel BDV antigen and RNA in the CNS of patients who presented with a history of mental disorders. BDV-specific antigen and RNA was also determined for the first time for the p16, p56 and L polymerase BDV-encoded polypeptides.
Thus, the present invention now unequivocally identifies the presence of infectious BDV in humans and its association with clinical profiles of mental disorders whose etiology remains unknown.